Program Project Grant: Bone Cell Structue and Gene Expression

描述(由申请人提供)：我们的计划由3个研究项目和2个核心组成，继续解决这样的假设，即核结构参数(染色质组织和核基质相关亚核转录位点的组装和活性)有助于基因表达，介导骨骼发育和动态平衡所需的骨细胞表型特性的开始、进展和维持。我们正在寻求一种综合的、多学科的团队方法，将分子、细胞、生化和体内遗传学方法结合起来，以确定核酸和调节蛋白的亚核组织促进生理信号整合的机制，以支持体内骨骼基因表达和骨发育的能力。我们的计划1)从结构和功能上(体外和体内)鉴定了将Runx2转录因子引导到调控转录的核内部位的核内运输信号；2)对染色质结构、核小体组织和染色质重塑与骨骼基因表达的功能相互关系进行了开创性的研究；3)证明Runx2为在靶基因启动子上的战略位置和调控蛋白阈值浓度动态配置功能复合体的核微环境中组装骨骼基因表达的组合成分提供了一个支架；4)通过促进调控网络的组织和活性，为理解调节骨骼基因表达的组合机制提供了一个新的维度；5)观察到在有丝分裂和向子代细胞分裂的过程中，骨表型特异性转录因子在靶基因上的保留，为决定细胞命运和谱系承诺提供了一种新的表观遗传学控制成分。我们将专注于进一步确定骨骼基因表达调控复合体的时空组织和组装，方法是确定依赖于亚核区域稳定复合体形成的目标基因，以及支持在有丝分裂过程中保持骨骼基因表达能力的机制。 成骨前体细胞(项目1)，核微环境中具有特殊共调节因子的多聚体复合体对成骨细胞的正向和负向调控(项目2)，以及以维生素D依赖的方式诱导和发展成骨细胞表型所需的染色质重塑(项目3)。相关性：核结构-基因表达相互关系的评估将为调节参数提供生物学验证，这些参数是骨骼发育所必需的，可能在代谢性骨骼疾病中受到影响。我们的研究将提供对核组织成分的见解，这些成分可以作为创新治疗的靶点。

DESCRIPTION (provided by applicant): Our program, configured as 3 research projects and 2 cores, continues to address the hypothesis that parameters of nuclear structure (chromatin organization and the assembly and activity of nuclear matrix-associated subnuclear sites for transcription) contribute to gene expression that mediates the onset, progression and maintenance of bone cell phenotypic properties required for skeletal development and homeostasis. We are pursuing an integrated, multidisciplinary team approach that combines molecular, cellular, biochemical and in vivo genetic approaches to define mechanisms by which subnuclear organization of nucleic acids and regulatory proteins facilitates integration of physiological signals to support competency for skeletal gene expression and bone development in vivo. Our program has 1) identified, structurally and functionally (in vitro and in vivo) characterized the intranuclear trafficking signal that directs the Runx2 transcription factor to sites within the nucleus where regulatory machinery governing transcription resides; 2) pioneered investigation of functional interrelationships of chromatin structure, nucleosome organization and chromatin remodeling with skeletal gene expression; 3) demonstrated that Runx2 provides a scaffold for assembly of combinatorial components of skeletal gene expression at strategic sites on target gene promoters and in nuclear microenvironments where threshold concentrations of regulatory proteins dynamically configure functional complexes; 4) provided a new dimension to understanding combinatorial mechanisms that mediate skeletal gene expression by facilitating the organization and activity of regulatory networks; and 5) observed the retention of bone phenotype-specific transcription factors at target genes during mitosis and partitioning to progeny cells providing a novel component of epigenetic control for cell fate determination and lineage commitment. We will focus on further defining spatial and temporal organization and assembly of regulatory complexes for skeletal gene expression by identifying target genes dependent on stable complex formation in subnuclear domains and mechanisms supporting retention of competency for skeletal gene expression during mitotic division of osteoprogenitor cells (project 1), the positive and negative regulation of osteoblastogenesis by multimeric complexes with specialized coregulatory factors in nuclear microenvironments (project 2), and the requirements for chromatin remodeling for induction and progression of the osteoblast phenotype in a vitamin D dependent manner (project 3). Relevance: evaluation of nuclear structure-gene expression interrelationships will provide biological validation of regulatory parameters that are obligatory for skeletal development and may be compromised in metabolic bone disease. Our studies will provide insights into components of nuclear organization that can be targeted for innovative therapy.